It has also been reported that extra-nuclear irradiation resembles biological effects of nuclear-traversed irradiation and production of free radical species is a key mediator of the process. Nuclear factor erythroid 2-related factor 2 (Nrf2) is an important regulator of cellular oxidative status. It remains unknown whether the Nrf2 will be activated in cells exposed to cytoplasm targeted irradiation. To this point, a study with normal human lung fibroblast WI38 on Single Particle Irradiation System to Cells (SPICE) facility at NIRS was performed. The results showed that cytoplasm targeted radiation with proton promoted the nuclear transfer of Nrf2. A dose-dependent manner of Nrf2 nuclear transfer was observed within the dose range from 0 to 1000 protons. By measuring the fraction of Nrf2 in nucleus, it was found that upon cytoplasm targeted with 500 protons, the accumulation of Nrf2 in nucleus enhanced with the extension of post-radiation time. The nuclear transfer of Nrf2 after cytoplasm targeted was controlled by activated MAPK ERK1/2, which was shown by the facts that the phosphorylation of MAPK ERK1/2 in WI38 increased after radiation, and the application of MAPK ERK1/2 inhibitor U0126 blocked the nuclear transfer of Nrf2. We further investigated the effect of activation of Nrf2 on radiation induced DNA damage and the results showed that pre-activation of Nrf2 with chemical inducer will alleviated DNA double strand breaks caused by cytoplasm targeted radiation. In conclusion, the current work indicated that Nrf2 antioxidative response was activated after cytoplasm targeted radiation and this might play a protective role against radiation induced DNA damage.The 12th International Workshop on Microbeam Probes of Cellular Radiation Response